Mast cell stabilizers for treatment of hypercytokinemia and viral infection

ABSTRACT

The present invention relates to methods for treating hypercytokinemia and viral infections associated with hypercytokinemia using a mast cell stabilizing compound, optionally in combination with an antiviral agent. The invention further relates to compositions and dosage forms comprising mast cell stabilizing agents, optionally with an antiviral agent.

STATEMENT OF PRIORITY

This application is a continuation of and claims priority to U.S. patentapplication Ser. No. 15/697,835, filed Sep. 7, 2017, which claims thebenefit of U.S. Provisional Application Ser. No. 62/385,021, filed Sep.8, 2016, the entire contents of each of which are incorporated byreference herein.

FIELD OF THE INVENTION

The present invention relates to methods for treating hypercytokinemiaand viral infections and other disorders associated withhypercytokinemia using a mast cell stabilizing compound, optionally incombination with an antiviral agent or other therapeutic agent. Theinvention further relates to compositions and dosage forms comprisingmast cell stabilizing agents, optionally with an antiviral agent orother therapeutic agent.

BACKGROUND OF THE INVENTION

Mast cells are a unique hematopoietic cell that is resident only intissue and not in the blood stream. They are sentinels, constantly onthe lookout for invading organisms, toxic molecules and tissue damage.When mast cells encounter such pathogens or damage, they release a floodof mediators; these mediators are a mix of small molecule effectors(such as histamine), proteases, lipid-derived signaling molecules(prostaglandins) and cytokines. The process of releasing these mediatorsis generically termed “degranulation.”

The release of mediators via degranulation results in the recruitment ofa wide variety of immune cells to the site of mediator release to fightthe offending intruder. This process also induces localized inflammationto restrict the movement of any infection or toxin. A controlledinflammatory response to foreign bodies enables the body to prevent thespread of toxins to neighboring tissues, limiting damage to one area andis desirable. Once the toxin has been neutralized, the normal course isfor the body to begin an active resolution of inflammation response(ROI). This results in an egress of recruited immune cells from theinjured tissue, tissue remodeling, a reduction in swelling and finallycomplete healing.

In some cases, this normal course of a proportional inflammatoryresponse, followed by resolution is not followed. This can result ineither a chronic state of inflammation or an overly robust mediatorresponse that is out of proportion to the invading event. The latterscenario has been termed a “cytokine storm” or hypercytokinemia. Anextremely high level of inflammatory cytokines and mediators that is notresolved is harmful to the host and can result in death, especially ifthe run-away response occurs in the lungs. The most damaging aspects andsymptoms of many diseases are a direct result of hypercytokinemia, asopposed to damage from the original disease.

Current treatments for conditions that induce hypercytokinemia do notgenerally do anything to ameliorate the ongoing production ofinflammatory mediators that perpetuate this condition. These conditionsare usually treated by interventions that target the pathogen inducingthe response or by using antagonists to one specific inflammatorymediator. While it is critical for the pathogens to be neutralized, inmost diseases that induce hypercytokinemia, the problem is not thepathogen, but rather the body's overly robust response to the pathogenby a dozen or more inflammatory mediators.

Nevertheless, down regulating the immune system is not enough on its ownto treat these conditions. In fact, there is evidence in some casessuggesting that treating patients with corticosteroids to reduceinflammation increases mortality. Corticosteroids are very powerful downregulators of the innate and adaptive immune system. “Turning off” allaspects of the immune response during an infection does not allow theclearance of the pathogen. Thus, a balance must be struck between anoverly robust immune response and too little of a response.

Influenza viruses are a related series of viruses in theOrthomyxoviridae family. Three types have been described (Types A, B orC) and all are negative-sense, segmented, single stranded RNA viruses.Types A and B are medically relevant to human health with type A beingthe cause of more severe disease. Type A viruses are normally found inwild birds and only rarely cause disease in their host. Influenza Asubtypes routinely cause waves of human influenza disease across theglobe with far reaching health and economic consequences. In the US,approximately 30,000 people die each year from flu or complicationsresulting from influenza infection. The estimates on economic impact inthe US are greater than $80 billion/year in direct medical costs andlost work. Influenza usually infects humans through the respiratorytract, although there are also reports that infection can occur throughthe eyes.

Influenza induces a rapid cytokine response in humans, usuallydetectable within hours of infection. An important point to note is thatthe severity of each case of flu is directly correlated with the levelsof induced cytokines. This is well known to those in the field and hasbeen documented across many different strains of influenza. What isparticularly striking about influenza induced disease is that severityis dependent on the viral strain and the individual. For instance, mostyearly cases of influenza are caused by the H1N1 strain. While manypeople are infected, the severity of the disease ranges from no symptomsto death. In a normal H1N1 seasonal flu season in the US, about 20% ofthe population are sickened (60 million people), 200,000 of those arehospitalized and about 30,000 people die each year (about 0.05%mortality). In outbreaks involving highly pathogenic strains of thevirus (H5N1 for instance) 30% to 60% of the cases are fatal. While thisindicates that the virus is the main cause of severity, the reason thatpeople exhibit more severe or fatal disease is the same with eitherstrain: the dangerously ill people have extremely high levels ofcytokines, regardless of viral strain. Thus, the problem is not reallythe virus itself, but rather the body's response to the virus. In morevirulent strains, a larger proportion of people will experiencehypercytokinemia. However, this hypercytokinemia is no different than itis in the small proportion of people that experience the same symptomsfrom a less virulent strain.

Mast cell stabilizers such as ketotifen and cromolyn (sodiumcromoglycate) have been shown to inhibit mast cell degranulation and theresulting release of mediators such as histamine, tumor necrosis factor(TNF)-α, prostaglandins, leukotrienes, interleukins and other cytokines.These effects may not be limited to mast cells and might have a broadereffect of reducing inflammatory cytokine release in multiple cell types.These compounds, however, do not impact the adaptive immune system andallow antibody based clearance of foreign bodies from the body tocontinue essentially as normal. Both of these compounds are used totreat chronic conditions. Cromolyn was discovered and used as an inhaledtreatment for asthma. Ketotifen was discovered as an H1 antihistamineand is used extensively in eye drop formulations to treat eyeinflammation. It was also developed as an oral treatment for asthma,although this seems to take several weeks for it to have significantimpact on chronic asthma. Ketotifen and to a lesser extent cromolyn havebeen shown to increase survival of mice in a model system of influenzainfection (Hu et al., Mast Cell-Induced Lung Injury in Mice Infectedwith H5N1 Influenza Virus, J. Virol. 86(6):3347 (2012); Han et al., Thetherapeutic effects of sodium cromoglycate against influenza A virusH5N1 in mice, Influenza and Other Respiratory Viruses 10(1):57 (2016)).In the case of ketotifen, it was combined with oseltamivir and theimpact on survival was increased when the compounds were combined.

The present invention overcomes shortcomings in the art by providingcompositions and methods for treating hypercytokinemia and disordersassociated therewith.

SUMMARY OF THE INVENTION

The present invention relates to the development of treatments to downregulate hypercytokinemia in order to intervene successfully in cases ofcytokine storm. The present invention provides a more effective way toaid the body in balancing inhibition of an overly robust immune responseand providing too little of a response to provide a treatment thatallows both problems to be addressed: the initial condition that thebody is reacting to (i.e., infection) and the resultinghypercytokinemia. This can be accomplished with a treatment regime thatdown regulates the innate immune system and allows the adaptive immunesystem to remove the offending pathogen. Alternatively, a treatment thatdown regulates the innate immune system can be combined with a treatmentto help clear the intruding particles (for example, an anti-viralcompound).

Accordingly, as one aspect, the invention provides a method of treating,ameliorating, or preventing hypercytokinemia in a subject in needthereof, comprising administering to the subject a therapeuticallyeffective or prevention effective amount of a mast cell stabilizingcompound, thereby treating, ameliorating, or preventing thehypercytokinemia. In some embodiments, the method further comprisesadministering to the subject an additional therapeutic agent, e.g., anantiviral agent.

The invention further relates to a method of treating, ameliorating, orpreventing a viral infection associated with hypercytokinemia in asubject in need thereof, comprising administering to the subject atherapeutically effective or prevention effective amount of a mast cellstabilizing compound, thereby treating, ameliorating, or preventing theviral infection. In some embodiments, the method further comprisesadministering to the subject an additional therapeutic agent, e.g., anantiviral agent.

The invention further relates to a method of treating, ameliorating, orpreventing a disorder associated with hypercytokinemia in a subject inneed thereof, comprising administering to the subject a therapeuticallyeffective or prevention effective amount of a mast cell stabilizingcompound, thereby treating, ameliorating, or preventing the disorder. Insome embodiments, the method further comprises administering to thesubject an additional therapeutic agent, e.g., an antiviral agent.

The invention further relates to a method of treating, ameliorating, orpreventing a fever in a subject in need thereof, comprisingadministering to the subject a therapeutically effective or preventioneffective amount of a mast cell stabilizing compound, thereby treating,ameliorating, or preventing the fever. In some embodiments, the methodfurther comprises administering to the subject an additional therapeuticagent, e.g., an antipyretic agent.

The invention also relates to a pharmaceutical composition comprisingnorketotifen and a pharmaceutically acceptable carrier, and optionallyan additional therapeutic agent.

The invention additionally relates to a pharmaceutical compositioncomprising ketotifen, an additional therapeutic agent, and apharmaceutically acceptable carrier.

The invention further relates to dosage forms comprising thepharmaceutical compositions of the invention.

The present invention is explained in greater detail in the drawingsherein and the specification set forth below.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows predicted serum levels of ketotifen in a subject duringseveral different dosing regimens.

DETAILED DESCRIPTION OF THE INVENTION

The present invention can be embodied in different forms and should notbe construed as limited to the embodiments set forth herein. Rather,these embodiments are provided so that this disclosure will be thoroughand complete, and will fully convey the scope of the invention to thoseskilled in the art. For example, features illustrated with respect toone embodiment can be incorporated into other embodiments, and featuresillustrated with respect to a particular embodiment can be deleted fromthat embodiment. In addition, numerous variations and additions to theembodiments suggested herein will be apparent to those skilled in theart in light of the instant disclosure, which do not depart from theinstant invention.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. The terminology used in thedescription of the invention herein is for the purpose of describingparticular embodiments only and is not intended to be limiting of theinvention.

Unless the context indicates otherwise, it is specifically intended thatthe various features of the invention described herein can be used inany combination.

Moreover, the present invention also contemplates that in someembodiments of the invention, any feature or combination of features setforth herein can be excluded or omitted.

To illustrate, if the specification states that a complex comprisescomponents A, B and C, it is specifically intended that any of A, B orC, or a combination thereof, can be omitted and disclaimed singularly orin any combination.

All publications, patent applications, patents, and other referencesmentioned herein are incorporated by reference herein in their entirety.

As used herein, “a,” “an,” or “the” can mean one or more than one. Forexample, “a” cell can mean a single cell or a multiplicity of cells.

Also as used herein, “and/or” refers to and encompasses any and allpossible combinations of one or more of the associated listed items, aswell as the lack of combinations when interpreted in the alternative(“or”).

Furthermore, the term “about,” as used herein when referring to ameasurable value such as an amount of a compound or agent of thisinvention, dose, time, temperature, and the like, is meant to encompassvariations of ±20%, ±10%, ±5%, ±1%, ±0.5%, or even ±0.1% of thespecified amount.

The term “consists essentially of” (and grammatical variants), asapplied to the compositions of this invention, means the composition cancontain additional components as long as the additional components donot materially alter the composition. The term “materially altered,” asapplied to a composition, refers to an increase or decrease in thetherapeutic effectiveness of the composition of at least about 20% ormore as compared to the effectiveness of a composition consisting of therecited components.

By the terms “treat,” “treating,” or “treatment of” (and grammaticalvariations thereof) it is meant that the severity of the subject'scondition is reduced, at least partially improved or stabilized and/orthat some alleviation, mitigation, decrease or stabilization in at leastone clinical symptom is achieved and/or there is a delay in theprogression of the disease or disorder.

The terms “prevent,” “preventing,” and “prevention” (and grammaticalvariations thereof) refer to prevention and/or delay of the onset of adisease, disorder and/or a clinical symptom(s) in a subject and/or areduction in the severity of the onset of the disease, disorder and/orclinical symptom(s) relative to what would occur in the absence of themethods of the invention. The prevention can be complete, e.g., thetotal absence of the disease, disorder and/or clinical symptom(s). Theprevention can also be partial, such that the occurrence of the disease,disorder and/or clinical symptom(s) in the subject and/or the severityof onset and/or the time period of symptoms is less than what wouldoccur in the absence of the present invention.

A “treatment effective” or “therapeutically effective” amount as usedherein is an amount that is sufficient to provide some improvement orbenefit to the subject. Alternatively stated, a “treatment effective”amount is an amount that will provide some alleviation, mitigation,decrease or stabilization in at least one clinical symptom in thesubject. Those skilled in the art will appreciate that the therapeuticeffects need not be complete or curative, as long as some benefit isprovided to the subject.

A “prevention effective” amount as used herein is an amount that issufficient to prevent and/or delay the onset of a disease, disorderand/or clinical symptoms in a subject and/or to reduce and/or delay theseverity of the onset of a disease, disorder and/or clinical symptoms ina subject relative to what would occur in the absence of the methods ofthe invention. Those skilled in the art will appreciate that the levelof prevention need not be complete, as long as some benefit is providedto the subject.

As used herein, the term “hypercytokinemia,” also known as a “cytokinestorm” or “cytokine cascade,” refers to an immune reaction consisting ofa positive feedback loop between cytokines and mediator secreting cellsresulting in highly elevated levels of various cytokines. Duringhypercytokinemia, the serum and/or organ levels of one or more cytokinemay be elevated, e.g., at least 5-fold higher than the level observed ina healthy subject or in a healthy population, e.g., at least 10-fold or100-fold higher. Symptoms of hypercytokinemia may include high fever,swelling and redness, extreme fatigue, and nausea.

A “disorder associated with hypercytokinemia” refers to any disorderthat has hypercytokinemia as one of its symptoms or side effects duringat least one stage of the disorder.

A “viral infection associated with hypercytokinemia” refers to any viralinfection that has hypercytokinemia as one of its symptoms or sideeffects during at least one stage of the infection.

“Influenza,” as used herein, is defined as an illness associated with apositive influenza diagnostic test.

“Influenza-like illness,” as used herein, is defined as the symptoms offever (temperature of 37.8° C. or higher), and a cough and/or a sorethroat without a known cause.

“Systemic inflammatory response syndrome (SIRS)” or “sepsis” is definedby the presence of two or more of the following clinical findings: (a)body temperature greater than 38° C. or 36° C.; (b) heart rate greaterthan 90 per minute; (c) respiratory rate greater than 20 per minute orPACO₂ less than 32 mmHg; (d) white blood cell count greater than 12,000cell/L or less than 4,000 cells/L, or greater than 10% immature (band)forms.

“Pharmaceutically acceptable,” as used herein, means a material that isnot biologically or otherwise undesirable, i.e., the material can beadministered to an individual along with the compositions of thisinvention, without causing substantial deleterious biological effects orinteracting in a deleterious manner with any of the other components ofthe composition in which it is contained. The material would naturallybe selected to minimize any degradation of the active ingredient and tominimize any adverse side effects in the subject, as would be well knownto one of skill in the art (see, e.g., Remington's PharmaceuticalScience; 21^(st) ed. 2005). Exemplary pharmaceutically acceptablecarriers for the compositions of this invention include, but are notlimited to, sterile pyrogen-free water and sterile pyrogen-freephysiological saline solution.

“Concurrently” means sufficiently close in time to produce a combinedeffect (that is, concurrently can be simultaneously, or it can be two ormore events occurring within a short time period before or after eachother). In some embodiments, the administration of two or more compounds“concurrently” means that the two compounds are administered closelyenough in time that the presence of one alters the biological effects ofthe other. The two compounds can be administered in the same ordifferent formulations or sequentially. Concurrent administration can becarried out by mixing the compounds prior to administration, or byadministering the compounds in two different formulations, for example,at the same point in time but at different anatomic sites or usingdifferent routes of administration.

The present invention is designed to address a physiological injury(incurred by any means) and the induced hypercytokinemia. This isaccomplished by treating the acute condition with a mast cellstabilizing compound that reduces the release of mediators from mastcells. These compounds may also be effective at reducing the release ofmediators from other cell types. The net effect is to reduce an overlyrobust reaction of the innate immune system, allowing the adaptiveimmune system to address the initial problem leading tohypercytokinemia. These compounds may also be combined with a diseasespecific-compound. The disease specific compound targets the initialinjury, but the injury may still have induced or continue to induce astrong cytokine response. The mast cell stabilizing compound reduces thereleased mediators, reducing hypercytokinemia. A reduction in the acuteinflammatory response prevents additional damage produced by out ofcontrol inflammation. A combined treatment approach is much moreefficacious than a single treatment because it addresses both theharmful injury (e.g., infection) and the even more deleterious cytokineresponse.

While some mast cell stabilizing compounds have been used in the past asgeneral anti-inflammatory agents, the present invention is distinct fromprior uses because it involves treating acute inflammatory conditions asopposed to chronic conditions (e.g., SIRS vs. asthma). In addition, thepresent invention involves treating symptoms caused by infectious agentssuch as viruses as opposed to allergic responses to seasonal allergens.

One of the most common conditions that can induce hypercytokinemia isinfluenza infection. Flu has a massive mortality and cost impact uponthe world each year, causing over 30,000 deaths a year on average in theUS. While there are vaccines for seasonal flu available, they havelimitations in that their effectiveness depends on choosing the correctstrains long before the flu season has arrived to enable time formanufacturing. Available antiviral therapies are also limited in theireffectiveness, because the virus is able to mutate and become resistantto all currently known antivirals. Even if the virus infecting a patientis sensitive to the antiviral used, inhibition of viral spread does nottreat a major health risk of influenza; the induced hypercytokinemia.

Mast cells are the resident sentinels that detect and react to aninitial infection by influenza virus. Viral recognition by mast cells isaccomplished by pattern recognition receptors and results in rapiddegranulation, releasing a plethora of cytokines (Graham et al., J.Immunol. 190: 4676 (2013)). This release of mediators calls in the firstresponders of the immune system, including T cells, to attack theinfection. Mast cells continue to release cytokines which spreadsystemically and continue to recruit more immune cells to the infection.These attracted cells also release cytokines to ramp up the immuneresponse. The result can be a self-reinforcing loop that escalates theresponse out of proportion to the infection. The acute symptoms that arefelt in flu are caused by the mediators, not the damage caused by thevirus. If left unchecked, this inflammatory response causes tissuedamage in the lungs and reduces lung function, while at the same timeproviding an ideal environment for bacterial growth that results inpneumonia. If left unchecked, the inflammatory response in the lungs canlead to acute respiratory distress syndrome (ARDS) and death.

Treating a patient with antivirals will not stop this self-reinforcingcycle. In fact, by the time a patient is experiencing symptoms, it is atleast 48 hours into an infection. Influenza viral budding begins 6 hoursafter a cell in infected and continues for days. By the time theinfection is detected and antiviral treatment has begun, there isalready a high viral load and a robust immune response: the patient isalready far down the road leading to cytokine storm. Thus, stopping thevirus alone at this stage is not enough to treat the physiologicalcondition of influenza-infected patients.

Thus, one aspect of the invention relates to a method of treating,ameliorating, or preventing hypercytokinemia in a subject in needthereof, comprising administering to the subject a therapeuticallyeffective or prevention effective amount of a mast cell stabilizingcompound, thereby treating, ameliorating, or preventing thehypercytokinemia. In some embodiments, the hypercytokinemia is due to adisease, disorder, or condition in the subject. In certain embodiments,the disease, disorder, or condition is not asthma, conjunctivitis, ormastocytosis. In some embodiments, the disease, disorder, or conditionis a viral infection, e.g., a respiratory virus infection, e.g., aninfluenza virus infection. In some embodiments, the disease, disorder,or condition is not influenza infection.

Another aspect of the invention relates to a method of treating,ameliorating, or preventing a disorder associated with hypercytokinemiain a subject in need thereof, comprising administering to the subject atherapeutically effective or prevention effective amount of a mast cellstabilizing compound, thereby treating, ameliorating, or preventing thedisorder.

Examples of clinical diagnoses that are associated with acutehypercytokinemia include, without limitation, viral infections such asinfluenza, influenza-like illness, systemic inflammatory responsesyndrome (from infectious or non-infectious causes), or sepsis.

In some embodiments, the methods are carried out on subjects that havebeen diagnosed with hypercytokinemia. In other embodiments, the methodsare carried out on subjects that have not been diagnosed withhypercytokinemia but have been diagnosed with or are suspected of havinga disease, disorder or condition associated with hypercytokinemia andfor which the present methods are expected to be beneficial. As can beseen by this list of clinical diagnoses, a documented elevation ofcytokines is not needed to identify patients that would benefit fromthis treatment. In fact, it is understood by those of skill in the artthat the detection of increased systemic levels of cytokines is notalways found in patients in which a cytokine storm is causing harmfuleffects. The local concentration of cytokines, particularly in vitalorgans, more specifically in the lungs, is of greater importance thanthe level of cytokines found in circulating plasma. Thus, from apractical perspective, relying on clinical diagnoses such as thoselisted above which do not measure cytokines but rather focus on symptomsor known causes of increased cytokine levels (like a positive influenzatest) are more effective in identifying patients that would benefit fromthe present invention.

Another aspect of the invention relates to a method of treating,ameliorating, or preventing a viral infection associated withhypercytokinemia in a subject in need thereof, comprising administeringto the subject a therapeutically effective or prevention effectiveamount of a mast cell stabilizing compound, thereby treating,ameliorating, or preventing the viral infection. In some embodiments,the viral infection is a respiratory virus infection, e.g., an influenzavirus infection.

In general, a primary viral infection or reactivated latent virusinduces a robust cytokine response in patients. This is particularlydangerous in respiratory viruses, but others can also induce dangerouslevels of cytokines. In these cases, reducing the magnitude of thehypercytokinemia is beneficial, regardless of which virus causes theresponse. Even in cases where the symptoms are not life-threatening, theelevation of cytokines causes symptoms and these can be ameliorated bytreatment with a mast cell stabilizing compound. Examples of viruseswhere this treatment method may be beneficial include, but are notlimited to, influenza, parainfluenza, norovirus, HIV, parvovirus,sindbis virus, dengue virus, adenovirus, Epstein-Barr virus (EBV),respiratory syncytial virus (RSV), vaccinia virus, metapneumovirus,rhinovirus, cytomegalovirus (CMV), hepatitis viruses (all types), herpessimplex virus (all types), papillomavirus, SARS coronavirus, MERScoronavirus, west Nile virus, yellow fever virus and others. In someembodiments, the virus is not influenza.

A further aspect of the invention relates to a method of treating,ameliorating, or preventing a fever in a subject in need thereof,comprising administering to the subject a therapeutically effective orprevention effective amount of a mast cell stabilizing compound, therebytreating, ameliorating, or preventing the fever. In some embodiments,the fever is associated with a respiratory virus infection, e.g., aninfluenza virus infection. In some embodiments, the fever is associatedwith a viral infection or other disorder associated withhypercytokinemia. In some embodiments, the antipyretic effect of themast cell stabilizing compound results in a decrease in body temperatureof at least 1° C., e.g., at 2° C., 3° C., 4° C., 5° C., or more.

In some embodiments, the methods of the invention further compriseadministering to the subject an additional therapeutic agent for thedisease, disorder, or condition. In some embodiments, the additionaltherapeutic agent is an antiviral agent, e.g., an anti-influenza agent.Examples of anti-influenza agents include, without limitation,rimantadine, amantadine, oseltamivir (Tamiflu®), laninamivir (Inavir®),zanamivir (Relenza®), peramivir (Rapivab®), tamiphosphor, favipiravir,VX-787, AL794, S-033188, AB103, TCAD Combo, Avi-7100, flufivirtide-3,fluconazole (FluCide®), ribavirin (Virazole), endonuclease inhibitors,matrix M1 inhibitors, antibodies to viral proteins, Vis-410,nitazoxanide, NT-300, fludase, alferon, PUR003, EV-077, surfaxin, andhomspera. In one embodiment, the anti-influenza agent is oseltamivir.Oseltamivir treatment for infected patients is dosed at 75 mg twice aday for 5 days. This provides a therapeutic dose systemically for aduration deemed appropriate for most cases of influenza A and B. In someembodiments, the additional therapeutic agent is an antipyretic agent,such as ibuprofen, naproxen, ketoprofen, nimesulide, aspirin, and othernon-steroidal anti-inflammatory drugs, and acetaminophen. In someembodiments, the additional therapeutic drug is an agent used to treatSIRS or sepsis, e.g., anti-anaphylaxis agents such as epinephrine,steroids, and diphenhydramine, or antioxidants such as selenium,glutamine, vitamin E, and eicosapentaenoic acid.

The mast cell stabilizing compound may be any mast cell stabilizingcompound known in the art. Examples of mast cell stabilizing compoundsinclude, without limitation, ketotifen, norketotifen, cromolyn,nedocromil, quercetin, pemirolast, olopatadine, ebastine andcarebastine. In some embodiments of the methods of the invention, themast cell stabilizing compound is ketotifen or a pharmaceuticallyacceptable salt thereof. In some embodiments of the methods of theinvention, the mast cell stabilizing compound is norketotifen or apharmaceutically acceptable salt thereof. In some embodiments, the mastcell stabilizing compound is a combination of ketotifen or apharmaceutically acceptable salt thereof and norketotifen or apharmaceutically acceptable salt thereof.

The mast cell stabilizing compound and the additional therapeutic agentmay be administered to the subject in the same composition or inseparate compositions, e.g., administered concurrently. The additionaltherapeutic agent can be delivered to the subject on a differentschedule or by a different route as compared to the mast cellstabilizing compound. The additional therapeutic agent can be any agentthat provides a benefit to the subject.

The methods of the present invention may also be combined with physicaltreatments that are known in the art to be effective for treating adisease, disorder, or condition. Physical treatments may include,without limitation, surgery (e.g., removal of tissue), transplantationof bone marrow or solid organs, plasmapheresis, modulation (increase ordecrease) of the temperature of a subject or a tissue or organ in thesubject, or any combination thereof.

The compounds of the invention may be administered to the subject asneeded to treat a disease, disorder, or condition. The compound can beadministered continuously or intermittently. In one embodiment, thecompound is administered to the subject more than once a day or onceevery 1, 2, 3, 4, 5, 6, or 7 days. In another embodiment, the compoundis administered to the subject no more than once a week, e.g., no morethan once every two weeks, once a month, once every two months, onceevery three months, once every four months, once every five months, onceevery six months, or longer. In a further embodiment, the compound isadministered using two or more different schedules, e.g., morefrequently initially (for example to build up to a certain level, e.g.,once a day or more) and then less frequently (e.g., once a week orless). In other embodiments, the compound can be administered by anydiscontinuous administration regimen. In one example, the compound canbe administered not more than once every three days, every four days,every five days, every six days, every seven days, every eight days,every nine days, or every ten days, or longer. The administration cancontinue for one, two, three, or four weeks or one, two, or threemonths, or longer. Optionally, after a period of rest, the compound canbe administered under the same or a different schedule. The period ofrest can be one, two, three, or four weeks, or longer, according to thepharmacodynamic effects of the compound on the subject.

An examination of peripheral cytokine levels in infected patients showsthat elevated circulating levels of cytokines, especially IP10 and IL-6,are present within hours of infection with a peak between day 2 and 4post-infection. Symptoms generally appear around 2 days post-infection.Cytokine levels rise to much higher levels in patients with more severesymptoms and remain elevated. Due to the variability of severity andcourse of infection from person to person, combination therapy may beadministered initially for 5 days and then continued until symptoms havesignificantly resolved towards baseline.

The compounds of the invention can be delivered to the subject by anysuitable route, e.g., oral, rectal, buccal (e.g., sub-lingual), vaginal,parenteral (e.g., subcutaneous, intramuscular, intradermal, orintravenous), topical (i.e., both skin and mucosal surfaces, includingairway surfaces), nasal and/or oral inhalation, and transdermaladministration.

Where the antiviral agent is administered by inhalation (e.g.,zanamivir), the mast cell stabilizing compound and the antiviral agentmay be combined in an inhaler for simultaneous delivery. The advantageof this approach is to deliver both active compounds directly to thesite of infection, potentially decreasing the time needed to reachtherapeutic levels in infected cells and thereby increasingeffectiveness. An additional advantage of using a mast cell stabilizingcompound (e.g., ketotifen with an inhaled antiviral) is itseffectiveness for treating asthma. One safety issue with zanamivir isthe appearance of adverse events in patients with asthma and chronicobstructive pulmonary disorder. It is possible that theanti-inflammatory properties of mast cell stabilizing compounds such asketotifen may reduce these events and increase the population in whichan inhaled antiviral can be utilized.

Some new antiviral therapies may be developed that require only a singledose. Two examples under development are an anti-flu monoclonal antibodyor a cap-dependent endonuclease inhibitor. In conjunction with suchtreatments, a mast cell stabilizing compound may be added to thetreatment and be delivered over several days, likely more than 3 days,in order to reduce the impact of hypercytokinemia. A time-releasemicrosphere approach could also be used to simultaneously deliver themast cell stabilizing compound and ensure that it is present for anextended time, even with only one dose. Shiny et al. (Int. J. Pharm.Investig. 3(3):119 (2013)) is an example of an approach for up to 30-daycontinuous delivery.

Some viral illnesses can cause severe symptoms requiringhospitalization. For patients in hospital it may be advantageous toprovide therapies by intravenous means. For this purpose, an antiviraland a mast cell stabilizing compound can be given through an IV,providing dosing of both medications for the required time period. Inthis way, seriously ill patients that are unable to swallow medicationwill be able to be treated with this combination of medication.

The compounds are delivered to the subject at a dose that is effectiveto treat the disorder. The effective dosage will depend on many factorsincluding the gender, age, weight, and general physical condition of thesubject, the severity of the disorder, the particular compound orcomposition being administered, the duration of the treatment, thenature of any concurrent treatment, the carrier used, and like factorswithin the knowledge and expertise of those skilled in the art. Asappropriate, a treatment effective amount in any individual case can bedetermined by one of ordinary skill in the art by reference to thepertinent texts and literature and/or by using routine experimentation(see, e.g., Remington, The Science and Practice of Pharmacy (21^(st) ed.2005)). In one embodiment, the mast cell stabilizing compound isadministered at a dose of about 0.001 to about 500 mg/kg body weight,e.g., about 0.001, 0.005, 0.01, 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7,0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 20, 30, 40, 50, 60, 70, 80, 90,100, 200, 300, 400, or 500 mg/kg or any range therein. In someinstances, the dose can be even lower, e.g., as low as 0.0005 or 0.0001mg/kg or lower. In some instances, the dose can be even higher, e.g., ashigh as 20, 50, 100, 500, or 1000 mg/kg or higher. For ketotifen thedosing range may be from about 0.001 mg/kg to about 40 mg/kg, e.g., fromabout 0.01 mg/kg to about 20 mg/kg, e.g., about 0.001, 0.005, 0.01,0.05, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 15, 20, 25, 30, 35, or 40mg/kg or any range therein. For norketotifen the dosing range may befrom about 0.001 mg/kg to about 500 mg/kg, e.g., from about 0.01 mg/kgto about 200 mg/kg e.g., about 0.001, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 2,3, 4, 5, 6, 7, 8, 9, 10 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70,75, 80, 85, 90, 95, or 100, 150, 200, 250, 300, 350, 400, 450, or 500mg/kg or any range therein. The present invention encompasses everysub-range within the cited ranges and amounts. These doses may beadministered once per day or more than once per day, e.g., 2, 3, or 4times per day.

Because symptoms of influenza do not generally appear until one or twodays into the infection, it is desirable that systemic therapeuticlevels of the mast cell stabilizing compound and the additionaltherapeutic agent be obtained quickly to prevent both viral spread andthe accumulation of higher level of cytokines. In order to accomplishthis in a way that will provide easy patient compliance, a time releaseform of administration may be advantageous. FIG. 1 illustrates serumlevels predicted for 4 mg per day ketotifen in several different dosingregimens based on published pharmacokinetic data.

Twice daily dosing at 2 mg per dose is the current approach used formast cell disorders. It takes at least 2.5 days to approach a steadystate level near 2 ng/ml. If the 4 mg/day dosing is taken in one dose,it surpasses the steady state level rapidly, but each day the overalllevel drops significantly and falls below 2 ng/ml. If a time releaseapproach is used with the first dose doubled to 8 mg, the 2 ng/ml levelis achieved quickly, the overall level does not go too high and aconsistent dose above 2 ng/ml is maintained. Thus, the use of a timerelease approach, optionally with an increased initial dose, allows fora rapid increase of systemic drug levels that are maintained throughoutthe treatment.

One property of many viruses, including influenza, is that they cause agreat deal of nasal inflammation and secretions. It may be advantageousto deliver this combination of an antiviral agent and mast cellstabilizing compound using an intranasal route (e.g., by nasalinstillation and/or nasal inhalation), providing the mast cellstabilizing compound directly to the sinus cavity. Sinus cavities arerich in vasculature, enabling many drugs to be efficiently absorbedsystemically using this approach. In fact, ketotifen has been shown tobe more bioavailable using intranasal delivery in rabbits than any otherroute. An additional benefit of a mast cell stabilizer like ketotifen isits antihistamine properties, allowing for a rapid and efficientreduction in severe nasal symptoms.

Some medications are delivered subcutaneously or intramuscularly toensure the availability of compounds that are not readily absorbed byother administration routes. This method also has the advantage of speedand being practical in hospitals, physician's offices, pharmacies andthe home. There are many auto injector systems available that wouldfacilitate self-injection of therapeutic doses of the combination of theinvention. In this case the compounds may be mixed in a single chamberif they are compatible, or a dual chamber system may be employed to keepthe antiviral and mast cell stabilizing compound in separatecompartments until dosing. As it is likely that the dosing schedule willbe several days, a system that allows for multiple dosing from a singledevice (or “pen”) would increase convenience and compliance with theneeded injection schedule.

For some patients (including children), swallowing tablets or pills isdifficult or impossible. In these cases, it may be advantageous todeliver the combination of the invention in an oral liquid formulation.This formulation may deliver the drug combination in a time releasemechanism or any other desired delivery profile. This may also beaccomplished using a suppository for intra-rectal delivery.

The present invention finds use in research as well as veterinary andmedical applications. Suitable subjects are generally mammaliansubjects. The term “mammal” as used herein includes, but is not limitedto, humans, non-human primates, cattle, sheep, goats, pigs, horses,cats, dog, rabbits, rodents (e.g., rats or mice), etc. Human subjectsinclude neonates, infants, juveniles, adults and geriatric subjects.

In particular embodiments, the subject is a human subject that hashypercytokinemia or a disorder associated with hypercytokinemia, e.g.,viral infections such as influenza, influenza-like illness, systemicinflammatory response syndrome, or sepsis, or a fever. In otherembodiments, the subject is a human subject at risk for developinghypercytokinemia or a disorder associated with hypercytokinemia, e.g., asubject that has been exposed to a virus or is living or working with asubject known to have a viral infection. When a virus like influenzainfects an individual, a detectable increase in systemic cytokinesstarts within hours of infection. While the associated localinflammation may serve to restrict spreading of the virus, in most casesthe systemic increases in cytokines does little to prevent the spread ofinfection and likely slows down the clearance of virus by the adaptiveimmune system. In addition, the accumulated fluids from inflammation aidin the spread of the virus through sneezing and coughing. Themicrodroplets produced enable viral particles to remain viable forextended times in an aerosol, allowing it to spread more effectively.The methods of the invention may be used prophylactically to reducesymptoms in anyone infected and reduce the spread of virus. In otherembodiments, the subject used in the methods of the invention is ananimal model of hypercytokinemia or a disorder associated withhypercytokinemia.

The subject can be a subject “in need of” the methods of the presentinvention, e.g., in need of the therapeutic and/or prophylactic effectsof the inventive methods.

In some embodiments, the subject is one that is in a higher riskpopulation for the viral infection, e.g., influenza infection. Thesewould include, but are not limited to, the elderly, the very young,pregnant women, immunocompromised patients, and ethnic groups shown tobe at higher risk (for example American Eskimo, Native American). Inaddition, very virulent strains of influenza have induced more severedisease in healthy individuals between the ages of 20 and 50 (thepandemic strain from 2009 being the most recent example), and thisregime is likely to be more effective in this group for a virulentstrain.

A further aspect of the invention relates to a pharmaceuticalcomposition comprising norketotifen and a pharmaceutically acceptablecarrier. The pharmaceutical composition may further comprise anadditional therapeutic agent, e.g., an antiviral agent, e.g., ananti-influenza agent, e.g., oseltamivir.

Another aspect of the invention relates to a pharmaceutical compositioncomprising ketotifen, an additional therapeutic agent, and apharmaceutically acceptable carrier. In some embodiments, the additionaltherapeutic agent is an antiviral agent, e.g., an anti-influenza agent,e.g., oseltamivir.

An additional aspect of the invention relates to a dosage formcomprising the pharmaceutical composition of the invention. The dosageform may be any dosage form known in the art that is suitable for themethods of the present invention. The dosage form may be, withoutlimitation, a solid or liquid oral dosage form, a dosage form for nasaland/or oral inhalation, a dosage form for intravenous administration, adosage form for transdermal or mucosal administration (e.g., a patch), adosage form for injection (e.g., subcutaneous or intramuscular), adosage form for implantation (e.g., a dissolvable formulation or adevice such as a pump), or a dosage form for ocular administration.

The compounds of this invention (e.g., mast cell stabilizing compoundssuch as ketotifen and norketotifen) include all pharmaceuticallyacceptable salt forms thereof. Examples of such salts include thosederived from pharmaceutically acceptable inorganic and organic acids andbases. Examples of suitable acid salts include, without limitation,acetate, adipate, alginate, aspartate, benzoate, butyrate, citrate,fumarate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride,hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate,malonate, methanesulfonate, nicotinate, nitrate, oxalate, palmoate,pectinate, persulfate, hydroxynapthoate, pivalate, propionate,salicylate, succinate, sulfate, tartrate, thiocyanate, tosylate andundecanoate. Other acids, such as oxalic, while not in themselvespharmaceutically acceptable, can be employed in the preparation of saltsuseful as intermediates in obtaining the compounds of the invention andtheir pharmaceutically acceptable acid addition salts.

Salts derived from appropriate bases include, without limitation, alkalimetal (e.g., sodium, potassium), alkaline earth metal (e.g., magnesiumand calcium), ammonium and N-(alkyl)₄ ⁺ salts.

Compounds of the invention include those having quaternization of anybasic nitrogen-containing group therein.

The discussion herein is, for simplicity, provided without reference tostereoisomerism. Those skilled in the art will appreciate that thecompounds of the invention can contain one or more asymmetric centersand thus occur as racemates and racemic mixtures, single opticalisomers, individual diastereomers, and diastereomeric mixtures. All suchisomeric forms of these compounds are expressly included in the presentinvention.

Similarly, compounds of the invention containing a double bond can existin the form of geometric isomers, which can be readily separated andrecovered by conventional procedures. Such isomeric forms are includedin the scope of this invention.

Unless otherwise stated, structures depicted herein are also meant toinclude compounds that differ only in the presence of one or moreisotopically enriched atoms. For example, compounds having the presentstructures except for the replacement of hydrogen by deuterium ortritium, or the replacement of a carbon by a ¹³C- or ¹⁴C-enriched carbonare within the scope of this invention. Such compounds are useful, forexample, as analytical tools or probes in biological assays.

Further, the compounds of the invention include prodrugs of thecompounds that are converted to the active compound in vivo. Forexample, the compound can be modified to enhance cellular permeability(e.g., by esterification of polar groups) and then converted by cellularenzymes to produce the active agent. Methods of masking charged orreactive moieties as a pro-drug are known by those skilled in the art(see, e.g., P. Korgsgaard-Larsen and H. Bundgaard, A Textbook of DrugDesign and Development, Reading U.K., Harwood Academic Publishers,1991).

The term “prodrug” refers to compounds that are rapidly transformed invivo to yield the parent compound of the above formula, for example, byhydrolysis in blood, see, e.g., T. Higuchi and V. Stella, Prodrugs asNovel delivery Systems, Vol. 14 of the A.C.S. Symposium Series and inEdward B. Roche, ed., Bioreversible Carriers in Drug Design, AmericanPharmaceutical Association and Pergamon Press, 1987, both of which areincorporated by reference herein. See also U.S. Pat. No. 6,680,299.Exemplary prodrugs include a prodrug that is metabolized in vivo by asubject to an active drug having an activity of the compounds asdescribed herein, wherein the prodrug is an ester of an alcohol orcarboxylic acid group, if such a group is present in the compound; anamide of an amine group or carboxylic acid group, if such groups arepresent in the compound; a urethane of an amine group, if such a groupis present in the compound; an acetal or ketal of an alcohol group, ifsuch a group is present in the compound; a N-Mannich base or an imine ofan amine group, if such a group is present in the compound; or a Schiffbase, oxime, acetal, enol ester, oxazolidine, or thiazolidine of acarbonyl group, if such a group is present in the compound, such asdescribed, for example, in U.S. Pat. Nos. 6,680,324 and 6,680,322.

The term “pharmaceutically acceptable prodrug” (and like terms) as usedherein refers to those prodrugs of the compounds of the presentinvention which are, within the scope of sound medical judgment,suitable for use in contact with the tissues of humans and/or otheranimals without undue toxicity, irritation, allergic response and thelike, commensurate with a reasonable risk/benefit ratio, and effectivefor their intended use, as well as the zwitterionic forms, wherepossible, of the compounds of the invention.

The compounds of the invention described above can be formulated foradministration in a pharmaceutical carrier in accordance with knowntechniques. See, e.g., Remington, The Science And Practice of Pharmacy(21^(st) ed. 2005). In the manufacture of a pharmaceutical compositionaccording to the invention, the compound is typically admixed with,inter alia, an acceptable carrier. The carrier must, of course, beacceptable in the sense of being compatible with any other ingredientsin the formulation and must not be deleterious to the patient. Thecarrier can be a solid or a liquid, or both, and can be formulated withthe compound as a unit-dose formulation, for example, a tablet, whichcan contain from 0.01% or 0.5% to 95% or 99% by weight of the compound.One or more compounds can be incorporated in the formulations of theinvention, which can be prepared by any of the well known techniques ofpharmacy comprising admixing the components, optionally including one ormore accessory ingredients.

The formulations of the invention include those suitable for oral,rectal, topical, buccal (e.g., sub-lingual), vaginal, parenteral (e.g.,subcutaneous, intramuscular, intradermal, or intravenous), topical(i.e., both skin and mucosal surfaces, including airway surfaces), nasaland/or oral inhalation, and transdermal administration, although themost suitable route in any given case will depend on the nature andseverity of the condition being treated and on the nature of theparticular active compound which is being used.

Formulations suitable for oral administration can be presented indiscrete units, such as capsules, cachets, lozenges, or tablets, eachcontaining a predetermined amount of the active compound; as amultiparticulate, powder or granules; as a solution or a suspension inan aqueous or non-aqueous liquid; or as an oil-in-water or water-in-oilemulsion. Such formulations can be prepared by any suitable method ofpharmacy which includes the step of bringing into association thecompound and a suitable carrier (which can contain one or more accessoryingredients as noted above). In general, the formulations of theinvention are prepared by uniformly and intimately admixing the compoundwith a liquid or finely divided solid carrier, or both, and then, ifnecessary, shaping the resulting mixture. For example, a tablet can beprepared by compressing or molding a powder or granules containing thecompound, optionally with one or more accessory ingredients. Compressedtablets can be prepared by compressing, in a suitable machine, thecompound in a free-flowing form, such as a powder or granules optionallymixed with a binder, lubricant, inert diluent, and/or surfaceactive/dispersing agent(s). Molded tablets can be made by molding, in asuitable machine, the powdered compound moistened with an inert liquidbinder.

The term “tablet” as used herein includes, but is not limited to,immediate release (IR) tablets, sustained release (SR) tablets, extendedrelease (ER) tablets, matrix tablets, multilayer tablets, multilayermatrix tablets, delayed release tablets and pulsed release tablets, anyor all of which may optionally be coated with one or more coatingmaterials, including polymer coating materials, such as entericcoatings, rate-controlling coatings, semi-permeable coatings and thelike. The term “tablet” also includes osmotic delivery systems in whicha drug compound is combined with an osmagent (and optionally otherexcipients) and coated with a semi-permeable membrane, thesemi-permeable membrane defining an orifice through which the drugcompound may be released. Tablet solid oral dosage forms that may beuseful in the practice of the invention include those selected from thegroup consisting of IR tablets, SR tablets, ER tablets, coated IRtablets, coated SR tablets, coated ER tablets, matrix tablets, coatedmatrix tablets, multilayer tablets, coated multilayer tablets,multilayer matrix tablets and coated multilayer matrix tablets. In someembodiments, a tablet dosage form is an enteric-coated tablet dosageform. In some embodiments, a tablet dosage form is an enteric-coatedextended release tablet dosage form.

The term “capsule” as used herein includes, but is not limited to, IRcapsules, SR capsules, ER capsules, coated IR capsules, coated SRcapsules, and ER capsules, including delayed release capsules. Capsulesmay be filled with powders, granules, multiparticulates, tablets,semi-solids, or liquids. In some embodiments, a capsule dosage form isan enteric-coated capsule dosage form. In some embodiments, a capsuledosage form is an enteric-coated extended release capsule dosage form.Capsules may be made of hard gelatin, soft gelatin, starch, cellulosepolymers, or other materials as known to the art.

The term “multiparticulate” as used herein means a plurality of discreteparticles, microparticles, nanoparticles, pellets, mini-tablets andmixtures or combinations thereof. If the oral form is a multiparticulatecapsule, hard or soft gelatin capsules or capsules of other materialscan suitably be used to contain the multiparticulate. In someembodiments, a sachet can suitably be used to contain themultiparticulate. In some embodiments, the multiparticulate may becoated with a layer containing rate controlling polymer material. Insome embodiments, a multiparticulate oral dosage form according to theinvention may comprise a blend of two or more populations of particles,pellets, or mini-tablets having different in vitro and/or in vivorelease characteristics. For example, a multiparticulate oral dosageform may comprise a blend of an instant release component and a delayedrelease component contained in a suitable capsule.

In some embodiments, the multiparticulate and one or more auxiliaryexcipient materials can be compressed into tablet form such as amultilayer tablet. In some embodiments, a multilayer tablet may comprisetwo layers containing the same or different levels of the same activeingredient having the same or different release characteristics. In someembodiments, a multilayer tablet may contain different activeingredients in each layer. Such a tablet, either single layered ormultilayered, can optionally be coated with a controlled release polymerso as to provide additional controlled release properties. In someembodiments, a multiparticulate dosage form comprises a capsulecontaining delayed release rapid onset minitablets. In some embodiments,a multiparticulate dosage form comprises a delayed release capsulecomprising instant release minitablets. In some embodiments, amultiparticulate dosage form comprises a capsule comprising delayedrelease granules. In some embodiments, a multiparticulate dosage formcomprises a delayed release capsule comprising instant release granules.

Formulations suitable for buccal (sub-lingual) administration includelozenges comprising the compound in a flavored base, usually sucrose andacacia or tragacanth; and pastilles comprising the compound in an inertbase such as gelatin and glycerin or sucrose and acacia.

Formulations of the present invention suitable for parenteraladministration comprise sterile aqueous and non-aqueous injectionsolutions of the compound, which preparations are preferably isotonicwith the blood of the intended recipient. These preparations can containanti-oxidants, buffers, bacteriostats and solutes which render theformulation isotonic with the blood of the intended recipient. Aqueousand non-aqueous sterile suspensions can include suspending agents andthickening agents. The formulations can be presented in unit/dose (e.g.,in a syringe or other injection device) or multi-dose containers, forexample sealed ampoules and vials, and can be stored in a freeze-dried(lyophilized) condition requiring only the addition of the sterileliquid carrier, for example, saline or water-for-injection immediatelyprior to use. Extemporaneous injection solutions and suspensions can beprepared from sterile powders, granules and tablets of the kindpreviously described. For example, in one aspect of the presentinvention, there is provided an injectable, stable, sterile compositioncomprising one or more compounds, in a unit dosage form in a sealedcontainer. The compound is provided in the form of a lyophilizate whichis capable of being reconstituted with a suitable pharmaceuticallyacceptable carrier to form a liquid composition suitable for injectionthereof into a subject. The unit dosage form typically comprises fromabout 0.001 mg to about 10 grams of the compound. When the compound issubstantially water-insoluble (e.g., when conjugated to a lipid), asufficient amount of emulsifying agent which is physiologicallyacceptable can be employed in sufficient quantity to emulsify thecompound in an aqueous carrier. One such useful emulsifying agent isphosphatidyl choline.

Formulations suitable for rectal administration are preferably presentedas unit dose suppositories. These can be prepared by admixing thecompound with one or more conventional solid carriers, for example,cocoa butter, and then shaping the resulting mixture.

Formulations suitable for topical application to the skin preferablytake the form of an ointment, cream, lotion, paste, gel, spray, aerosol,or oil. Carriers which can be used include petroleum jelly, lanoline,polyethylene glycols, alcohols, transdermal enhancers, and combinationsof two or more thereof.

Formulations suitable for transdermal administration can be presented asdiscrete patches adapted to remain in intimate contact with theepidermis of the recipient for a prolonged period of time. Formulationssuitable for transdermal administration can also be delivered byiontophoresis (see, for example, Pharm. Res. 3:318 (1986)) and typicallytake the form of an optionally buffered aqueous solution of thecompound. Suitable formulations comprise citrate or bis\tris buffer (pH6) or ethanol/water and contain from 0.1 to 0.2 M active ingredient.

Other pharmaceutical compositions can be prepared from the compoundsdisclosed herein, such as aqueous base emulsions. In such an instance,the composition will contain a sufficient amount of pharmaceuticallyacceptable emulsifying agent to emulsify the desired amount of thecompound. Particularly useful emulsifying agents include phosphatidylcholines and lecithin.

In addition to compound, the pharmaceutical compositions can containother additives, such as pH-adjusting additives. In particular, usefulpH-adjusting agents include acids, such as hydrochloric acid, bases orbuffers, such as sodium lactate, sodium acetate, sodium phosphate,sodium citrate, sodium borate, or sodium gluconate. Further, thecompositions can contain microbial preservatives. Useful microbialpreservatives include methylparaben, propylparaben, and benzyl alcohol.The microbial preservative is typically employed when the formulation isplaced in a vial designed for multidose use. Other additives that arewell known in the art include, e.g., detackifiers, anti-foaming agents,antioxidants (e.g., ascorbyl palmitate, butyl hydroxy anisole (BHA),butyl hydroxy toluene (BHT) and tocopherols, e.g., α-tocopherol (vitaminE)), preservatives, chelating agents (e.g., EDTA and/or EGTA),viscomodulators, tonicifiers (e.g., a sugar such as sucrose, lactose,and/or mannitol), flavorants, colorants, odorants, opacifiers,suspending agents, binders, fillers, plasticizers, lubricants, andmixtures thereof. The amounts of such additives can be readilydetermined by one skilled in the art, according to the particularproperties desired.

The additive can also comprise a thickening agent. Suitable thickeningagents can be those known and employed in the art, including, e.g.,pharmaceutically acceptable polymeric materials and inorganic thickeningagents. Exemplary thickening agents for use in the presentpharmaceutical compositions include polyacrylate and polyacrylateco-polymer resins, for example poly-acrylic acid and poly-acrylicacid/methacrylic acid resins; celluloses and cellulose derivativesincluding: alkyl celluloses, e.g., methyl-, ethyl- andpropyl-celluloses; hydroxyalkyl-celluloses, e.g.,hydroxypropyl-celluloses and hydroxypropylalkyl-celluloses such ashydroxypropyl-methyl-celluloses; acylated celluloses, e.g.,cellulose-acetates, cellulose-acetatephthallates,cellulose-acetatesuccinates and hydroxypropylmethyl-cellulosephthallates; and salts thereof such as sodium-carboxymethyl-celluloses;polyvinylpyrrolidones, including for example poly-N-vinylpyrrolidonesand vinylpyrrolidone co-polymers such as vinylpyrrolidone-vinylacetateco-polymers; polyvinyl resins, e.g., including polyvinylacetates andalcohols, as well as other polymeric materials including gum traganth,gum arabicum, alginates, e.g., alginic acid, and salts thereof, e.g.,sodium alginates; and inorganic thickening agents such as atapulgite,bentonite and silicates including hydrophilic silicon dioxide products,e.g., alkylated (for example methylated) silica gels, in particularcolloidal silicon dioxide products. Such thickening agents as describedabove can be included, e.g., to provide a sustained release effect.However, where oral administration is intended, the use of thickeningagents as aforesaid will generally not be required and is generally lesspreferred. Use of thickening agents is, on the other hand, indicated,e.g., where topical application is foreseen.

Further, the present invention provides liposomal formulations of thecompounds disclosed herein. The technology for forming liposomalsuspensions is well known in the art. When the compound is in the formof an aqueous-soluble material, using conventional liposome technology,the same can be incorporated into lipid vesicles. In such an instance,due to the water solubility of the compound, the compound will besubstantially entrained within the hydrophilic center or core of theliposomes. The lipid layer employed can be of any conventionalcomposition and can either contain cholesterol or can becholesterol-free. When the compound of interest is water-insoluble,again employing conventional liposome formation technology, the compoundcan be substantially entrained within the hydrophobic lipid bilayerwhich forms the structure of the liposome. In either instance, theliposomes which are produced can be reduced in size, as through the useof standard sonication and homogenization techniques. The liposomalformulations containing the compound disclosed herein, can belyophilized to produce a lyophilizate which can be reconstituted with apharmaceutically acceptable carrier, such as water, to regenerate aliposomal suspension.

In certain embodiments, the dosage form is an extended release dosageform. The dosage form may have a half-life of at least 24 hours. In someembodiments, the extended release dosage form will provide a slowerrelease of the drug over a 24-hour time period such that a consistent,tight range of the drug is available systemically for 24 hours. Incertain embodiments, as shown in the example in FIG. 1, the serum levelof the compound is kept above 2 ng/mL throughout the treatment period,but at the same time does not rise above 2.5 ng/mL. The dosage form maycomprise and/or be coated with a rate controlling polymer material,e.g., hydroxypropyl methylcellulose, a polymer of acrylic or methacrylicacid or their respective esters, or copolymers of acrylic or methacrylicacid or their respective esters.

In any of the above-mentioned embodiments, a controlled release coating(e.g., an enteric coating) may be applied to the final dosage form(capsule, tablet, multilayer tablet, etc.). The controlled releasecoating may typically comprise a rate controlling polymer material asdefined above. The dissolution characteristics of such a coatingmaterial may be pH dependent or independent of pH.

The term “rate controlling polymer material” as used herein includeshydrophilic polymers, hydrophobic polymers and mixtures of hydrophilicand/or hydrophobic polymers that are capable of controlling or retardingthe release of the compound from a dosage form of the present invention.Suitable rate controlling polymer materials include those selected fromthe group consisting of hydroxyalkyl cellulose such as hydroxypropylcellulose and hydroxypropyl methyl cellulose; poly(ethylene) oxide;alkyl cellulose such as ethyl cellulose and methyl cellulose;carboxymethyl cellulose; hydrophilic cellulose derivatives; polyethyleneglycol; polyvinylpyrrolidone; cellulose acetate; cellulose acetatebutyrate; cellulose acetate phthalate; cellulose acetate trimellitate;polyvinyl acetate phthalate; hydroxypropylmethyl cellulose phthalate;hydroxypropylmethyl cellulose acetate succinate; polyvinylacetaldiethylamino acetate; poly(alkylmethacrylate) and poly (vinylacetate). Other suitable hydrophobic polymers include polymers and/orcopolymers derived from acrylic or methacrylic acid and their respectiveesters, zein, waxes, shellac and hydrogenated vegetable oils.Particularly useful in the practice of the present invention are polyacrylic acid, poly acrylate, poly methacrylic acid and poly methacrylatepolymers. Some of these polymers (e.g., poly methacrylate polymers) canbe used as delayed release polymers to control the site where the drugis released.

The foregoing is illustrative of the present invention, and is not to beconstrued as limiting thereof. The invention is defined by the followingclaims, with equivalents of the claims to be included therein. Allpublications, patent applications, patents, patent publications, and anyother references cited herein are incorporated by reference in theirentireties for the teachings relevant to the sentence and/or paragraphin which the reference is presented.

1. A method of treating or ameliorating hypercytokinemia in a subject inneed thereof, comprising administering to the subject a therapeuticallyeffective amount of norketotifen or a pharmaceutically acceptable saltthereof, thereby treating or ameliorating the hypercytokinemia, whereinthe hypercytokinemia is caused by a systemic inflammatory response. 2-5.(canceled)
 6. The method of claim 1, further comprising administering tothe subject an additional therapeutic agent for the systemicinflammatory response. 7-9. (canceled)
 10. A method of treating orameliorating a systemic inflammatory response associated withhypercytokinemia in a subject in need thereof, comprising administeringto the subject a therapeutically effective amount of norketotifen or apharmaceutically acceptable salt thereof, thereby treating orameliorating the systemic inflammatory response associated withhypercytokinemia. 11-12. (canceled)
 13. The method of claim 10, furthercomprising administering to the subject an additional therapeutic agentfor the systemic inflammatory response. 14-23. (canceled)
 24. The methodof claim 1, further comprising performing a physical treatment on thesubject.
 25. The method of claim 24, wherein the physical treatment issurgery, transplantation, plasmapheresis, modulation of temperature, orany combination thereof. 26-43. (canceled)
 44. The method of claim 6,wherein the norketotifen or a pharmaceutically acceptable salt thereofand the additional therapeutic agent are administered in the samecomposition.
 45. The method of claim 6, wherein the norketotifen or apharmaceutically acceptable salt thereof and the additional therapeuticagent are administered in separate compositions.
 46. The method of claim13, wherein the norketotifen or a pharmaceutically acceptable saltthereof and the additional therapeutic agent arc administered in thesame composition.
 47. The method of claim 13, wherein the norketotifenor a pharmaceutically acceptable salt thereof and the additionaltherapeutic agent are administered in separate compositions.